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ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613
Citation: ZHANG Zhan, FENG Yan, LI Qian, CUI Li. Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii[J]. Journal of China Pharmaceutical University, 2021, 52(6): 742-750. DOI: 10.11665/j.issn.1000-5048.20210613
shu

Characterization of a UDP-glucose 4-epimerase from Acinetobacter Baumannii

  • 1.

    State Key Laboratory of Microbial Metabolism, School of Life Science and Technology, Shanghai Jiao Tong University, Shanghai 200240

  • 2.

    School of Life Science & Technology, China Pharmaceutical University, Nanjing 210009, China

Funds: This study was supported by the National Natural Science Foundation of China (No.21977067, No.31770098, No.31620103901) and the National Key R&D Program of China (No.2018YFA0900702)
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  • Received Date: April 21, 2021
  • Revised Date: November 03, 2021
    Graphical Abstract
    • Abstract
  • The purpose of this article is to express the UDP-glucose 4-epimerase from Acinetobacter baumannii AB0057, characterize its enzymatic properties and analyze its structure and function.The epimerase gene was constructed into pET-28a expression vector and heterologously expressed in BL21(DE3).Enzyme activity was assayed using high performance liquid chromatography.The structure and key residues were analyzed by phylogenetic analysis, sequence alignment, homology modeling and molecular docking.Results indicated that the recombinant enzyme Gne1 was expressed at a molecular weight of 38.9 kD, with an optimum temperature of 44 °C and an optimum pH of 6.0 .Michaelis-Menten parameters KM and kcat were (1.227 ± 0.082 4) mmol/L and (82.64 ± 3.562) × 10-3?min-1, respectively.This enzyme belongs to NADB_Rossmann superfamily and UDP_G4E_1_SDR_e subgroup with typical GXGXXG and YXXXK sequence motifs.The N-terminal structural domain bound to NAD, while the C-terminal structural domain bound to substrate, and the catalytic key sites were S125 and Y150.The current work verified the epimerase activity of Gne1, explained its sequence and structural features, revealed its binding mode with substrates and cofactors, and analyzed the key residues, which provides a basis for protein engineering to improve the epimerase activity and then use biological enzymatic method to synthesize rare functional sugars.
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